The Use of Pharmaceutical and Non-pharmaceutical Grade Compound in Animals and Labeling Expectations

IACUC Policy #045-05, Approved 12/14/2012, Last Revised: 05/17/2024

Background

Federal regulations require Investigators to use pharmaceutical-grade compounds for injection in animals, even in acute procedures including euthanasia. This includes, but is not limited to, medications/drugs, vehicles, and diluents. Pharmaceutical-grade compounds meet established standards of purity and composition helping ensure animal health and experimental results.    

It is recognized that many experimental compounds used in research are not available as pharmaceutical grade, or that pharmaceutical grade compounds may need to be diluted or combined for use in laboratory animal research. The purpose of this policy is to outline the additional justification and details that need to be captured within the animal use protocol when non-pharmaceutical grade or compounded agents are used. In addition, this policy outlines the labeling and storage requirements for compounded agents. The use of chemical grade compounds, compounded drugs, or dilution of drugs can introduce unexpected or even toxic effects and should be avoided when possible.  

Definitions

• Pharmaceutical Grade Compounds: are any active or inactive drug, biologic, reagent, etc., manufactured under Good Manufacturing Practices, which is approved, conditionally approved, or indexed by the U.S. Food and Drug Administration (FDA) or for which a chemical purity standard has been written or established by a recognized compendia, such as the United States Pharmacopeia – National Formulary (USP-NF) and the British Pharmacopoeia (BP). USP-NF labeling does not equate to pharmaceutical grade.
• Non-Pharmaceutical Grade (NPG): compounds are chemicals or compounds that do not meet the criteria noted above in respect to pharmaceutical grade compounds. This would include pure chemicals obtained from any chemical supplier that require reconstitution. Phosphate buffered saline (PBS) is also considered non-pharmaceutical as it is not intended for parenteral injection.
• The United States Pharmacopeia National Formulary (USP-NF): provides FDA-enforceable quality standards for drugs, dietary supplements, and excipients as well as procedures for tests, assays and analytical methods. Parenteral articles meet Pharmacopeia requirements for sterility, pyrogens, particulate matter, and other contaminants, and, where appropriate, contain inhibitors of the growth of microorganisms.
• An injection: is a preparation intended for administration through the skin or directly into a blood vessel, organ, tissue, or lesion.
• Compounding: is a practice in which a person combines, mixes, or alters ingredients of a drug to create a medication tailored to the needs of an individual patient(s).

Requirements

All agents used in animals must be listed in the IACUC protocol. In addition, the use of non- pharmaceutical-grade compounds must be clearly identified in the animal use protocol and approved by the IACUC. In the protocol, investigators should indicate:

• Justification for use (i.e., non-availability of an acceptable veterinary or human pharmaceutical-grade compound or formulation or scientific necessity).
• Processes that will be used to ensure purity and sterility. 
• Cost savings is not a justification for using non-pharmaceutical-grade compounds, although OLAW and USDA have made exceptions in cases of limited access resulting in exorbitant costs.

If compounding of drugs to be injected in animals is necessary, investigators must ensure:

• Aseptic preparation (compounding) of the drug including preparation and storage in sterile vials and filtration or autoclaving of compounded drug if original components are not sterile. Injectable drugs should never be used if they contain particulate matter, precipitates, turbidity, or discoloration.
• Appropriate storage of the compounded drug to include the use of a secondary container and methods which maintain sterility yet allows repeat draws e.g. use of a sterile injection vial with a rubber stopper. Alternatively, for single-use purposes, a sterile microfuge tube can be used.
• Appropriate labeling of containers for storage of compounded drugs to include:
  • Name of the compound(s) and diluent (when applicable)
  • Final concentration (usually mg/ml)
  • Date of Expiration
    • Mixtures/Dilutions: the earliest expiration date listed on the stock bottles of agents used.
    • Experimental (NPG) compounds:  the expiration date should be based on performance evaluation of the agent(s) for efficacy as well as consideration of the frequency of use/method of storage.
  • IACUC approval for the use of Non-pharmaceutical grade compounds is based on:
    • Scientific necessity
    • Non-availability of an acceptable veterinary or human pharmaceutical-grade compound or formulation
    • Appropriate process for aseptic preparation of compounded drugs for injection in animals
    • Cost savings is not a justification for using non-pharmaceutical-grade compounds, although Office of Laboratory Animal Welfare (OLAW) and U.S. Department of Agriculture (USDA) have made exceptions in cases of limited access resulting in exorbitant costs.

Note: It is highly recommended when using compounded drugs that a lab-specific SOP be created to establish a proper preparation procedure and expiration for the compounded drug. The length of time a compounded drug can be used should be based on use frequency and performance. Compounded drugs may result in decreased “shelf-life” compared to the stock drug(s). Furthermore, the potential for bacterial contamination is increased when compounding drugs or when drawing from multi-use vials. 

References

• FAQs about the PHS Policy on humane care and use of laboratory animals. Wolff A, et al. Lab Animal (NY). 2003 Oct; 32(9):33-6. 
• Animal Research Advisory Committee Guidelines, NIH, OLAW  
• OLAW: Educational Resources: Use of Non- Pharmaceutical-Grade Chemicals and Other Substances in Research with Animals   
• The United State Pharmopeia National Formulary  http://library.ohio-state.edu/record=b5907458~S7  
• The Orange Book is the reference for FDA-approved human drugs.
• The Green Book is the reference for FDA-approved veterinary drugs.

Approved SOP for Use of Tribromoethanol (Avertin)

Background

Tribromoethanol (TBE), commonly referred to by the brand name Avertin, has been used as an anesthetic agent for mice. TBE provides rapid induction and recovery for single use, short duration (approximately 15-20 minutes) surgical procedures in rodents. Improper preparation, storage, or use of Tribromoethanol can result in severe side effects including peritonitis, ileus, and death, particularly when repeated doses are administered or ineffective anesthesia. Specifically, TBE degrades in the presence of heat and light, producing toxic byproducts that are potent gastrointestinal irritants. TBE is not available as a pharmaceutical-grade compound and alternative options for rodent anesthesia including both pharmaceutical-grade injectable and inhalation options are readily available. Regulatory agencies have subsequently recommended that the use of TBE be discontinued as an anesthetic in mice when possible. ULAR veterinary staff is available to discuss alternatives to the use of TBE.  

Use

The body of scientific literature on this compound, as well as the regulatory requirement to use pharmaceutical-grade drugs in animals, presents a compelling case for discontinuing the use of TBE. The IACUC will review the continued use of TBE at the time of three-year renewal on a case-by-case basis; researchers will be required to provide justification for the continued use of this agent for survival surgery. 

Storage and Expiration Period

TBE must be stored at 2-8°C in light-protected containers. When stored at this temperature, the solution may be used for up to two weeks.  Frozen TBE solution can be stored at -80°C for up to 6 months.

The animal use protocol can reference this SOP or alternatively provide details regarding the preparation of the working solution and proper storage of the stock solution in the animal use protocol.

References

• Papaioannou, VE and Gox, JG. Efficacy of Tribromoethanol Anesthesia in Mice. Laboratory Animal Science, 1993. April, 43(2): 189-192.
• Zeller, WM; Burki, G; and Panoussis, B. Adverse Effects of Tribromoethanol as Used in the Production of Transgenic Mice. Laboratory Animal Science, 1998. October, 32(4): 407-413.
• Kohn, DF; Wixson, SK; White, WJ; and Benson, GJ. Anesthesia and Analgesia in Laboratory Animals, 1997.
• Lieggi, C.C., et al., Efficacy and safety of stored and newly prepared tribromoethanol in ICR mice. Contemp Top Lab Anim Sci, 2005. 44(1): 17-22.
• PHS Policy on the Human Care and Use of Laboratory Animals, Frequently Asked Questions
• Lieggi, C.C., et al., An evaluation of preparation methods and storage conditions of tribromoethanol. Contemp Top Lab Anim Sci, 2005. 44(1): 11-6.
• Meyer, R.E. and R.E. Fish, A review of tribromoethanol anesthesia for production of genetically engineered mice and rats. Lab Anim (NY), 2005. 34(10): 47-52.
• Zeller, W., et al., Adverse effects of tribromoethanol as used in the production of transgenic mice. Lab Anim, 1998. 32(4): p 407-13.
• Koizumi, T., H. Maeda, and K. Hioki, Sleep-time variation for ethanol and the hypnotic drugs tribromoethanol, urethane, pentobarbital, and propofol within outbred ICR mice. Exp Anim, 2002. 51(2): p119-24. National Human Genome Research Institute Guideline 03.2.

Approved SOP for Use of Saturated Potassium Chloride (KCl)

Background

The American Veterinary Medical Association (AVMA) Guidelines for the Euthanasia of Animals includes intravenous or intracardiac injection of a solution of supersaturated potassium chloride into an animal under general anesthesia as an acceptable method to enact a quick and humane death consistent with adequate veterinary practices. This method of euthanasia is preferable when euthanizing livestock or wildlife species to reduce environmental exposures and the risk of toxicosis for predators or scavengers in situations where carcasses of euthanized animals may be consumed. Saturated KCl is only available as a non-pharmaceutical-grade compound.

Use

The potassium ion is cardiotoxic, and rapid intravenous or intracardiac administration of 1 to 2 mmol/kg of body weight will cause cardiac arrest. Saturated potassium chloride has been commonly used to euthanize large animals (rabbits, dogs, cats, sheep, pigs, etc.) under general anesthesia as approved in the AVMA Guidelines for the Euthanasia of Animals. Saturated potassium chloride is not available in a pharmaceutical grade form. Scientific justification for the use of this euthanasia method is to accomplish a quick and humane death during a non-survival procedure in an animal that is under general anesthesia and this agent does not contaminate the animal carcass per environmental landfill requirements.

References

• AVMA [American Veterinary Medical Association]. 2020. Guidelines for the Euthanasia of Animals. Schaumburg, IL: AVMA.
• U.S. Department of Agriculture, Animal and Plant Health Inspection Service, Animal Care, Policy 3- Veterinary Care, April 14, 1997.
• Barbiturates. In: Ciganovich E, ed. Field manual of wildlife diseases. US Department of the Interior/US Geological Survey, Biological Resources Division, Information and Technical Report 199-2001.
• Lumb W. Euthanasia by noninhalent pharmacologic agents. J Am Vet Med Association. 1974; 165: 851-852.